The Role of NhaP1, NhaP2, and NhaP3 Antiporters in the Acid Tolerance of Vibrio cholerae

Vibrio cholerae (Vc), the etiological agent of cholera, has developed many mechanisms to combat the great fluctuations in pH, ion content and osmotic conditions of the vastly differing environments it encounters through its infections of the human host. A series of cation antiporters are found in the Vc membrane to assist the organism to combat these varying conditions. An estimated twelve antiporters reside in the bacterial membrane including NhaP1, NhaP2, and NhaP3. These three proteins have been identified as K+(Na+)/H+ antiporters specifically active in low pH conditions. We tested the hypothesis that these three antiporters are required for the survival of Vibrio cholerae in low pH environments. In this study the “6x6 drop plate method” by Chen et al. (10) was used to compare the survival of Vibrio cholerae O395N1 (wild type) to Vc-O395N1 ∆nhaP1∆nhaP2∆nhaP3 (the triple mutant) in acidic conditions over a period of 90 minutes. At 15 minutes in pH 3.5 potassium-modified Luria broth, the wild type had a statistically significant higher percentage of survival than the triple mutant in comparison to non-acid treated cells (2.08% compared to 0.0223%, P<0.0001). When comparing complemented nhaP1, nhaP2, or nhaP3 to the wild type strain, complemented nhaP3 was able to restore the wild type phenotype (P=0.1251) while complemented nhaP1 or nhaP2 showed reduced survivability/culturability at 15 minutes at pH 3.5 (P=0.0003 and P